This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The development of cutting edge technologies for imaging the mitochondrial metabolic function including the redox state of biological tissues are in great need, since mitochondria and its functional/genetic abnormalities have been connected to a number of diseases including cancer, diabetes, cardiovascular diseases, and neurodegenerative diseases. Low temperature NADH/Fp (reduced nicotinamide adenine dinucleotide/oxidized flavoproteins) fluorescence imaging or "redox scanning" has been developed since about 30 years ago to image the in vivo mitochondrial redox states of tissues on the basis of the redox ratios (Fp/NADH or Fp/(NADH+Fp)). A CCD-based redox imager has also been built to acquire images much faster with higher spatial resolution (~10 pm). However, redox ratio is based on the ratio of the relative intensities of the fluorescence signals of NADH and Fp, which usually depend on instrument settings such as filters and lamp conditions. In this project we aim to develop a calibration procedure to quantify NADH and Fp signals so that redox images obtained at different time or with different settings can be compared. We also aim to upgrade the hardware and software of the old redox scanner to a new pc-control unit that will acquire redox images faster with higher SNR and signal dynamic ranges. We also aim to develop tissue imaging methods for reactive oxygen species, which are connected to mitochondrial redox state and also implicated to play key role in the generation and progression of many diseases.